Carnitine and its use in reducing cardiac toxicity and as a synergist with cytostats

ABSTRACT

Carnitine may be administered for the purpose of reducing cardiac toxicity resulting from administration of a cytostat.

CROSS-REFERENCE

This is a division of Ser. No. 917,944 filed June 22, 1978, which is adivisional of Ser. No. 799,473 filed May 23, 1977 which is a CIP of Ser.No. 681,063 filed Mar. 28, 1976 which is CIP of Ser. No. 485,301 filedJuly 2, 1974, U.S. Pat. No. 3,968,241, which is CIP of Ser. No. 303,772filed Nov. 6, 1972 U.S. Pat. No. 3,830,931.

In the above patents, Carnitine, which isβ-hydroxy-γ-trimethyl-aminobutyric acid, and its pharmaceuticallyacceptable salts are shown to be useful for improving myocardialcontractility and systolic rhythmn in congestive heart failure and shockin humans and are also shown to be useful in treating cardiacarrhythmias. According to my copending application, Ser. No. 681,063,Carnitine is useful for improving the myocardial function of a human oranimal heart when the oxygen supply to such heart has been diminished,which may occur during heart failure or during surgery which maydirectly or indirectly involve the heart.

Carnitine per se is known and is sold in Europe as an appetitestimulant. It has been reported that the material has an effect on thegrowth rate of children; see E. G. Bornich et al., Clinic Chemica Acta,5, 1960, and Alexander et al., "Protides in the Biological Fluids", 6thColloquium, Bruges, 1958, pp 306-310. Investigation of the drug as anantagonist of thyroid hormone in cases of hyperthyroidism has also beenreported, Gilgore et al., Journal of New Drugs, 6, No. 6, 319, 320(1966), and DeFelice et al., loc. cit., 6, No. 6, 351, 353 (1966). Ihave also disclosed the use of Carnitine in the treatment of anginapectoris in my copending application Ser. No. 57,741 filed June 28,1971, now abandoned.

While the mechanism of action of β-hydroxy-γ-trimethylaminobutyric acidin these various conditions is not fully understood, there is noimmediately apparent reason to expect from these previous uses thatadministration of carnitine or a pharmaceutically acceptable saltthereof would be useful for other purposes.

β-hydroxy-γ-trimethylaminobutyric acid demonstrates a positive inotropicaction on the myocardium, which contractile force is not associated withan abnormal decrease in heart rate. In addition to restoring the sinusrhythmn to normal, β-hydroxy-trimethylaminobutyric acid results in asignificant increase in cardiac output. Dramatic diureses with reversalof the hypotension and urine retention which can accompany congestiveheart failure is often seen, together with a restoration of pulse andblood pressure. Significantly, the toxicity ofβ-hydroxy-γ-trimethylaminobutyric acid is remarkably low. The drug canthus be given orally or, more significantly parenterally, such asintravenously, without concern to toxic reactions. For improving themyocardial function, carnitine and its pharmaceutically acceptable saltsis administered parenterally.

Its properties can be seen in both animal models and clinical studies.

Thus in the isolated perfused Langendorf heart preparation,β-hydroxy-γ-trimethylaminobutyric acid effectively minimizes thereduction in the force of myocardial contraction during periods ofreduced blood flow. This protection lasts for at least 60 minutes withno adverse effect on the vital functions of the heart itself. This maybe seen from the following.

Twenty adult mongrel dogs weighing from 10-12 kg and of either sex wereanesthetized with sodium pentobarbital, 30 mg/kg. A polyethylenecatheter was placed in the right femoral artery to allow for the rapidremoval of blood. Clotting was prevented by injecting 1000 units/kgheparin (sodium) into the femoral vein. The chest was opened by amidline incision and all major branches of the ascending aorta, exceptthe brachiocephalic trunk, were ligated. The reservoir chamber of theperfusion system was filled with autologous blood drawn from thecatheterized femoral artery of the same dog. The brachiocephalic arterywas then cannulated with a short stainless steel cannula which wasconnected by polyethylene tubing to the pump perfusion circuit. At thistime the descending aorta was ligated and the heart quickly removed fromthe chest cavity. The heart was then suspended in the heated reservoirchamber and perfused with blood by means of a Harvard peristalsic pump.The blood was maintained at 35° C. throughout and was oxygenated bybubbling 95% oxygen and 5% carbon dioxide into the reservoir chamber.Flow rates were found to vary slightly from heart preparation to heartpreparation and ranged from 150 to 250 ml/min. Perfusion pressure wasmaintained at 25 to 50 mm Hg and was used as a direct index of coronaryvascular resistance. R=(P/F) Flow=constant. A needle-tipped catheterconnected to a Sanborn strain gauge and recorder was used tocontinuously monitor the perfusion pressure. Needle-tipped electrodeswere placed into the ventricles of the heart to monitorelectrocardiogram and heart rate. Force of contraction was measured viaa Walton-Brodie strain gauge arch sutured to the left ventricle.

Following a 30-minute equilibration period the blood flow to the heartwas reduced by 50%. Five minutes after the reduction of coronary flowthe change in force of contraction was measured. This reduction in forceis expressed as percent change from control. In most preparations thisprocedure was repeated twice prior to the administration ofβ-hydroxy-γ-trimethylaminobutyric acid. The dose ofβ-hydroxy-γ-trimethylaminobutyric acid varied from 50 to 250 mg, and wasinjected directly into the inflow side of the perfusion circuit leadingdirectly into the heart. At from 5 to 60 minutes after injection ofβ-hydroxy-γ-trimethylaminobutyric acid, the coronary blood flow wasagain reduced by 50% and the change in force measured. Only those heartsin which repeated measurement could be obtained were included in thestudy.

In each of the 20 isolated perfused hearts, a reduction in coronary flowproduced a significant decrease in force of contraction. Percentdecrease prior to administration of β-hydroxy-γ-trimethylaminobutyricacid varied from 55% to 85% with a mean of 71±5. One hundred mg ofβ-hydroxy-γ-trimethylaminobutyric acid limited the percent decrease inforce of contraction to from 19.6±3.4 to 28.4±7.9. At a dose of 150 mg,the precent decrease in force was from 25.0±7.2 to 29.5±9.7. Followingthe administration of 200 mg, the decrease in force of contractionfollowing a 50% reduction in blood flow was from 25.0±6.4 to 27.3±4.1.There is a significant difference between the control decrease in forceof contraction and those noted following administration ofβ-hydroxy-γ-trimethylaminobutyric acid. In no instance was there anoverlap between control and treated measurements of reduced flow ±standard deviation.

Restriction of blood flow to the dog myocardium by partial ligation ofthe anterior descending coronary artery is a useful model to study theeffects of drugs in ischemic myocardial disease. In untreated (control)dogs, restricting coronary blood flow by 50% resulted in markedelectrocardiographic changes including ST-segment elevation, which is adiagnostic feature of acute myocardial ischemia. Reducing coronary bloodflow by 75% produced ventricular fibrillation.

Six dogs weighing about 10-12 kg were treated with DL-carnitine (100mg/kg, intravenously). Five minutes after drug administration theanterior descending coronary artery was occluded to reduce blood flow by50%. In contrast to the six untreated animals, the dogs that weretreated with carnitine had only minor alterations in electrocardiograms.Carnitine treatment resulted in a 90% reduction in ST-segment elevationcompared to control dogs. When blood flow was reduced by 75%, carnitinetreatment resulted in a 50% to 60% decrease in ST-segment elevationcompared to control dogs. In addition, carnitine protected the heartsagainst ventricular fibrillation.

β-hydroxy-γ-trimethylaminobutyric acid also demonstrates a significanteffect on failing heart-lung preparation, as can be seen from thefollowing.

A series of 10 adult mongrel dogs were anesthetized with pentobarbitalsodium, 30 mg/kg, and the chest of each opened via midline incision. Theascending aorta was cannulated by inserting a large-bore polyethylenecatheter into the brachiocephalic artery and advancing it into theaortic circulation. The descending aorta and the subclavian artery werethen ligated following injection of heparin sodium, 1000 units/kg. Theaortic catheter was connected to a heated blood bath into which allcardiac output was pumped. The bath was also connected to a second largecatheter which was inserted into the superior vena cava. The azygos veinand the inferior cava were ligated. A bilaterial vagotomy was performed.Aortic blood flow, heart rate, venous pressure, reservoir level and leftventricular force of contraction were continuously monitored. Resistanceto aortic flow was then increased by 200% for 30 minutes, causing aprogressive decrease in flow, heart rate and force of contraction. At 30minutes, resistance to flow was decreased from 200% to 100% of controlvalues. No significant recovery of force, rate or flow was noted inuntreated heart-lung preparations. Those preparations given 250 mg/kg ofβ-hydroxy-γ-trimethylaminobutyric acid just prior to decreasing theresistance to flow at 30 minutes demonstrated a 50% to 75% of force ofcontraction, heart rate and flow rates. These changes persisted for 30to 60 minutes or until termination of the experiment (usually at 1 to11/2 hours).

The effect of β-hydroxy-γ-trimethylaminobutyric acid on atrial flutterand ventricular arrhythmias associated with congestive heart failure anda comparison with the corresponding effect of Quabain can be seen fromthe following:

Twenty adult mongrel dogs, anesthetized with sodium pentobarbital, 30mg/kg, were placed in severe heart failure using a method described byGattschalk. Electrical stimuli were applied to the atrial appendages ofthe heart to induce atrial flutter and ventricular arrhythmias.Following 30 minutes of decreased cardiac work and ventriculararrhythmias, the dogs were given either Quabain (1 mg/kg) orβ-hydroxy-γ-trimethylaminobutyric acid (20 mg/kg). Quabain produced a40%±10% increase in the force of contraction, a 20%±5% decrease in theheart rate, and a 32%±5% increase in the cardiac output. Overall cardiacwork increased 20%±2%. No anti-arrhythmiac effects were observed. Thedogs treated with β-hydroxy-γ-trimethylaminobutyric acid showed an85%±35% increase in force of contraction, a 40%±10% increase in heartrate, and a 90%±12% increase in cardiac output. Cardiac work increased60% and 15%, and in 8 to 10 dogs, the arrhythmias were converted to anormal sinus rhythm.

The anti-arrhythmic effect noted above can be more clearly seen indaunomycin-induced arrhythmias in both the Langendorff preparation andin intact Rhesus monkeys.

In a series of 20 isolated perfused Landendorff preparations, daunomycinwas administered in a dose of 100 mg/kg thereby consistently producingventricular arrhythmias, tachycardia, and severe arrhythmias. Following30 minutes of the severe arrhythmias, β-hydroxy-γ-trimethylaminobutyricacid was administered in doses of from 50 mg to 500 mg. In 18 of the 20isolated heart preparations, the arrhythmias noted were eliminated. Thearrhythmias which were eliminated did not reappear for periods up to 60to 90 minutes.

Twelve intact monkeys were likewise given daunomycin at a dose of 100mg/kg. After 15 to 30 minutes of observed arrhythimas,β-hydroxy-γ-trimethylaminobutyric acid was administered intravenously ina dose of 100 to 250 mg. In 9 of the 12 intact monkeys preparations, thearrhythmias were eliminated. The monkeys were followed for from 8 to 12hours using continuous recording techniques. No further arrhythmias werenoted. No toxic manifestation was noted and all of the monkeys uponsubsequent autopsy gave negative pathological reports.

The observations noted above in various laboratory models were confirmedin clinical studies which are briefly summarized below.

CASE 1

A patient with carcinoma of the colon had both ureters implanted intohis colon. Before surgery he was hypertensive and had a left branchblock. After surgery, his blood pressure fell to 90/40.β-hydroxy-γ-trimethylaminobutyric acid was given as a single IV dose of500 mg and within 60 seconds the blood pressure was 160/90. The cardiacrate fell from 105 to 85. The patient put out 615 cc of urine within anhour after β-hydroxy-γ-trimethylaminobutyric acid was given, 400 cc ofwhich were excreted within the first 30 minutes. In the previous ninehours there was a total urine output of less than 400 cc. His QRScomplex on the EKG averaged 0.11 before admininstration and two minutesafter administration averaged around 0.08-9. A few days later his bloodpressure dropped from the previous consistent level of approximately160/80 to 130/70. He was then given another dose of 500 mg IV and within60 seconds his blood pressure rose to 180/80.

CASE 2

A similar type of operation, i.e. carcinoma of the colon and bilateralimplantation of ureters, had been performed. The patient suddenly becamehypotensive and entered the intensive care unit. He had a cardiac rateof 125 and the blood pressure was not detectable. A single IV dose of500 mg of β-hydroxy-γ-trimethylaminobutyric acid was given. In 15minutes, the systolic pressure was 80. Total urinary output for thetwenty-four hours preceding this dose ofβ-hydroxy-γ-trimethylaminobutyric acid had 175 cc. The urinary outputfor 24 hours following β-hydroxy-γ-trimethylaminobutyric acidadministration was 970 cc, a substantial portion of which occurredwithin an hour. The patient had atrial fibrillation before theadministration of the drug which disappeared within 30 seconds. Thenormal sinus rhythm which occurred was maintained.

CASE 3

The patient, age 50, had a myocardial infarct with a history ofhypertension. He went into cardiogenic shock and his blood pressure wasnot measurable. He responded to norepinephrine and his blood pressurerose to 85/65. He developed a gallop rhythm. He was given 500 mg ofβ-hydroxy-γ-trimethylaminobutyric acid in two doses thirty minutesapart. His blood pressure rose to 100/70 and his urinary output rose to160 cc/hour (compared with a previous output of 10 cc/hour). The galloprhythm remained.

CASE 4

A 63 year old male was hospitalized repeatedly for cardiac failure dueto congestive heart failure. He had been treated for 14 years withcardiac glycosides and had intermittent cardiac irregularities. Uponthis admission, the patient was found to be in cardiac failure withsevere and continuing arrhythmias and no detectable blood pressuredespite a continuous infusion of levophed. The patient was then given500 mg of β-hydroxy-γ-trimethylaminobutyric acid. An immediate recoveryof normal sinus rhythm, an improved cardiac output and a detectableblood pressure within the normal range were noted. Within one hour afteradministration, the patient, who had previously been anuric for 61/2hours, began to put out significant quantities of urine. This improvedcondition persisted for 24 hours. The patient received no furthertreatment with the drug.

CASE 5

A 42 year old male with a history of cardiovascular difficulties rangingfrom early hypertension to mitral valve replacement was admitted withcongestive heart failure. The patient had previously been treated withcoronary vasodilators and/or cardiac glycosides. Following 28 hours ofsupportive therapy, to which he was non-responsive, the patient wasgiven 500 mg of β-hydroxy-γ-trimethylaminobutyric acid every four hours.The clinical response of the patient was evident within a few hours andhe was essentially asymptomatic. After seven days he was discharged fromthe hospital. β-hydroxy-γ-trimethylaminobutyric acid reversed both thecardiac arrhythmias and the congestive heart failure in this patient.

CASE 6

A 62 year old male suffered a myocardial infarct but demonstrated noacute cardiovascular difficulties. β-hydroxy-γ-trimethylaminobutyricacid was, nevertheless, given, in a dose of 500 mg, to determine whetherthere would be any cardiovascular effect. As expected, no detectablecardiovascular effect was observed over a period of two hours.

CASE 7

A 58 year old male was admitted with an acute myocardial infarct. He hadno great difficulty with his blood pressure but had prematureventricular contractions as noted on the EKG. An intravenous dose of 500mg of β-hydroxy-γ-trimethylaminobutyric acid was given and within a fewminutes his PVC's were greatly diminished. A second dose was given tenminutes after the first dose and within a few minutes the PVC'scompletely disappeared. There was no noticeable effect on bloodpressure.

CASE 8

A 73 year old male patient with a large myocardial infarct entered thehospital in cardiogenic shock. Doses of 500 mg ofβ-hydroxy-γ-trimethylaminobutyric acid were given at 30-minuteintervals. No detectable cardiovascular effect was observed.

CASE 9

A 65 year old male was admitted to the hospital with severe congestiveheart failure with marked pulmonary and peripheral edema. He was given500 mg of β-hydroxy-γ-trimethylaminobutyric acid every 15 minutes forover a period of approximately one hour, for a total of four doses. Adiuresis immediately followed and within a period of four hours hisclinical condition was markedly improved and his dyspnea virtuallydisappeared.

CASE 10

A 78 year old female was scheduled for a cholecystectomy. Her pulse was55 (left bundle branch block) and the anesthesiologist would not handlethe patient until her pulse was higher. Atropine was given without aresponse. An automatic cardiac pacemaker was inserted and the patient'spulse rate went to 80. After surgery the patient did well until thepacemaker was turned off. Her pulse immediately fell to 58-60.β-hydroxy-γ-trimethylaminobutyric acid was administered and her pulseimmediately went to 76. This same phenomena occurred twice with the nexthour; i.e. pulse fell and the drug reversed the fall. After the secondadministration, the patient was once again placed on the pacemaker. Whenthe pacemaker was turned off the pulse immediately fell. The third doseof β-hydroxy-γ-trimethylaminobutyric acid was then given and the pulseimmediately rose. From that point on the pacemaker was not required tomaintain a regular pulse that was 70 or over.

CASE 11

A 63 year old female was operated upon to correct a small bowel fistula.She had cancer of the bladder and had undergone bladder irradiation.Postoperatively she had excessive vaginal bleeding that was emanatingfrom the bladder. Her blood pressure began to fall and her pulse rateincreased. Eventually, her blood pressure was undetectable and her pulsefell to 60. β-hydroxy-γ-trimethylaminobutyric acid, 500 mg, was givenand almost immediately her systolic pressure rose to 90 and her pulse to100-120. In approximately one hour, her blood pressure was againundetectable and her pulse rate was 50. Once more 500 mg ofβ-hydroxy-γ-trimethylaminobutyric acid was given and duringβ-hydroxy-γ-trimethylaminobutyric acid administration her systolic bloodpressure rose to 90 and her pulse to 100. Additional administration ofthe drug was not made.

CASE 12

An 86 year old male was admitted with a diagnosis of severe anemia andcongestive heart failure. He was found to have carcinoma of the colon.He was digitalized and given 5 pints of blood whereupon he was operatedupon. Following surgery his EKG showed many ectopic beats and a bundlebranch block. Extrasystoles became quite prominent. Blood pressure was134/80 and pulse 58. He was given 500 mg of 62-hydroxy-γ-trimethylaminobutyric acid intravenously over a 5 minuteperiod and before drug administration was completed, the cardiacarrhythmias were markedly reduced. He had previously failed to respondto atropine. Three subsequent doses were given over a period of an hourwhereupon his arrhythmia disappeared except for an occasional ectopicbeat. His P-R interval shortened to a first degree heart block. A normalsinus rhythm continued throughout his hospital stay. Digitalis wasstopped before surgery.

Arrhythmias induced by the experimental removal of a heart from ananesthesized intact dog preparatory to the Langendorff perfusiontechnique occur in approximately 50 percent of the hearts isolated.These arrhythmias are varied and are both atrial (in 20 cases) andventricular (in 16 cases) in nature. In twenty arrhythmic isolated doghearts carnitine (100 mg, parenteral single dose) eliminated allarrhythmias and restored normal sinus rhythm.

As part of the open-chest experimental procedure hearts of anesthesizeddogs were exposed via a midline chest incision. The atrium of the heartwas electrically stimulated to cause atrial flutter and fibrillationsubsequent ventricular arrhythmias in 20 such preparations, Carnitinewhen administered intravenously in doses of 50-250 mg, eliminated theatrial and ventricular arrhythmias occurring in all preparations.

12 cats were similarly treated by electrical stimulations of the atriumof the heart to cause atrial flutter and fibrillation. Administration of20 mg/kg of carnitine i.v. produced a slight anti-arrhythmic effect.Administration of 50 mg/kg of carnitine i.v. returned the rhythms tonormal. Administration of 100 mg/kg of carnitine i.v. also returned therhythms to normal and resulted in a longer lasting effect than 50 mg/kg.

Twenty isolated dogs' hearts treated with 100 mg/kg of daunomycinresulted in severe atrial and ventricular arrhythmias in allpreparations. When carnitine was administered in a single dose of100-250 mg parenteral, the arrhythmias were eliminated in 18 of 20preparations.

Intact monkeys: daunomycin (100 mg/kg) caused severe atrial andventricular arrhythmias in twelve intact monkeys. In 9 of the 12 animalsthe severe arrhythmias were completely eliminated by carnitine (singledose of 100-250 mg i.v.).

Isolated dog hearts: Adriamycin, an analogue of daunomycin, exerts anarrhythmic effect similar to that induced by daunomycin. In sixangendorff heart preparations, all hearts were protected from thearrhythmic effect of 100 mg/kg adriamycin by carnitine administration(100-250 mg parenteral).

β-hydroxy-γ-trimethylaminobutyric acid contains a center of asymmetryand thus exists in two sterioisomers. Either the racemate or theindividual isomers can be employed. While the racemate can beconveniently employed, it appears the L-isomer is more active while theD-isomer is slightly more toxic. Thus the LD₅₀ in the mouse is asfollows:

    ______________________________________                                                   Subcutaneous                                                                              Intraperitoneal                                        ______________________________________                                        Racemate     11.5 g/kg     12 g/kg                                            L-isomer     13.8          14                                                 D-isomer     10.5          10                                                 ______________________________________                                    

The test data set forth above is illustrative of the use of carnitineand its pharmaceutically acceptable nontoxic salts for improving themyocardial function of a human or animal heart to which the oxygensupply has been diminished by parenterally administering to a human oranimal whose heart's oxygen supply has been diminished an amount ofactive agent sufficient to improve the myocardial function. Thepreferred method of parenteral administration is intravenous. Preferablyeither carnitine or the hydrochloride salt thereof is administered.

Fluid unit dosage forms for parenteral administration are prepared bysuspending or dissolving a measured amount ofβ-hydroxy-γ-trimethylaminobutyric acid in a non-toxic liquid vehiclesuitable for injection, such as an aqueous or oleagenous medium, andsterilizing the suspension or solution. Alternatively a measured amountof β-hydroxy-γ-trimethylaminobutyric acid is placed in a vial and thevial and its contents are sterilized and sealed. An accompanying vial orvehicle can be provided for mixing prior to administration.

Of the above fluid unit dosage forms intended for injection, inparticular intravenous and intramuscular injection, are preferred. Thesterile liquid or fluid injectible unit dosage forms are thecompositions of choice.

In addition to the free base, which can exist in Zwitterionic form, onecan employ pharmaceutically acceptable salts such as the hydrochloride(m.p. 142° ). Since, however, β-hydroxy-γ-trimethylaminobutyric acid asthe free base is itself soluble in aqueous media, utilization of a saltfor solubility purposes is generally not necessary.

The dose which is administered must of course be determined from theage, weight and condition of the patient, utilizing sound professionaljudgement. An improvement in the myocardial function can be observed atdoses as low as 1 to 2 mg/kg but generally the dose is from about 5 toabout 10 mg/kg. Larger doses can be given with relative safety in viewof the low toxicity of the compound and such doses are often indicatedwhen an insufficient response indicates an insufficient initial dose forthe particular patient. Utilization of oral unit dosage forms alsorequires a somewhat larger dose than is employed in the case ofparenteral administration.

EXAMPLE

A sterile aqueous suspension for intramuscular injection, containing ineach ml. 100 mg of β-hydroxy-γ-trimethylaminobutyric acid, is preparedfrom the following ingredients:

    ______________________________________                                        hydroxy-γ-trimethylaminobutyric acid                                                               gm 100                                             Sodium carboxymethylcellulose, low viscosity                                                             gm 10                                              Polysorbate 80, U.S.P.     gm 4                                               Propylparaben, U.S.P.      gm 0.4                                             Water for injection q.s.   gm 1,000                                           ______________________________________                                    

The present invention is concerned with two further novel aspects of theuse of Carnitine. The first of these is the reduction of cardiactoxicity by administering Carnitine to humans suffering cardiac toxicitycaused by the administration of cytostats. As set forth above, bothdaunomycin and adriamycin were utilized in animals to producearrhythmias for the purpose of demonstrating the use of Carnitine in thetreatment of arrhythmias. Similarly, as described above, adriamycin wasadministered to animals to exert an arrhythmic effect similar to thatinduced by daunomycin for the purpose of demonstrating the use ofCarnitine in the treatment of arrhythmias.

Daunomycin and adriamycin are representative of the following group ofcytostats:

N-acetyl-daunomycin

Danorubicin

Rubidazone

Adriamycin-14-octanoate

4'-epi-Adriamycin

Adriamycin, B-anomer

4'-epi-Adriamycin, B-anomer

Actinomycin

Carminomycin

Daunomycin oxine

These compounds are useful for their cytostatic activity in humand andanimals. When these compounds are administered for their cytostaticactivity, one frequent side effect is a toxic effect on the heart. Thus,according to one aspect of the present invention, Carnitine can beadministered for the purpose of reducing the cardiac toxicity resultingfrom administration of these cytostats to humans and animals for thepurpose of arresting and/or reducing the size of the tumors.

Thus, when Carnitine was administered following the daunomycin inducedarrhythmias, cardiac toxicity was significantly reduced or eliminated.The following data demonstrates a synergistic effect when Carnitine isadministered in synergistic combination with adriamycin:

MATERIALS AND METHODS

Cell and Culture Techniques. The Chinese hamster ovary (CHO) cell stockswere maintained in monolayer cultures in Hsu's modified McCoy's Medium5A, supplemented with 20% fetal calf serum in a 5% CO₂ -95% air,humidified incubator at 37° C. For experiments on exponentially growingpopulations, 10⁶ cells were placed into replicate 60-×15-mm Petri dishescontaining medium and 20% fetal calf serum, at least 18 hr beforetreatment.

Survival Determinations. The effects of Carnitine (100 ug/ml) onAdria-induced cell killing were determined on exponentially growingcells. Concentrations of from 1 to 10 μg Adria per ml were used.Replicate plates were used in each survival study and each experimentwas performed at least 3 times. The average of the survival fractions ateach dose point were plotted in the graph. In all cases survival wasdetermined by the ability of the treated cells to form colonies. Afterthe treatments, the cell monolayers were washed twice with Puck'sSolution A and trypsinized (0.025% for 5 min.), and known numbers ofsingle cells were plated into Petri dishes and incubated for 6 to 8 daysfor colony formation. Colonies were stained and counted. A cell wasconsidered to have retained reproductive capacity (viability) if it gaverise to a colony of 50 or more cells.

RESULTS

A continuous 3 hour exposure dividing cells to 100 ug/ml of Carnitinealone did not produce any cell-killing effect. The survival curve foradriamycin-treated cells was biphasic. 50% of the cells were killed by a1-hour exposure to as little as 0.5 ug/ml of adriamycin. Survivaldecreased exponentially as the dosage was increased to 2 ug/ml where theslope of the curve resumed a more resistant shape.

According to one embodiment of the present invention, treatment of thecells with Carnitine after a 1-hour exposure to adriamycin showeddramatic synergy in cytostatic activity.

What is claimed is:
 1. A method of reducing cardiac toxicity whichresults from the administration to a human of a cytostat selected fromthe group consisting of Rubidazone, Actinomycin and Carminomycin, whichresults in cardiac toxicity, which method comprises administering tosuch a human in need there of an amount ofβ-hydroxy-γ-trimethylaminobutyric acid, an isomer thereof, apharmaceutically acceptable salt thereof, or a pharmaceuticallyacceptable salt of an isomer thereof, sufficient to reduce cardiactoxicity.
 2. A method of reducing cardiac toxicity which results fromthe administration to a human of Rubidazone which results in cardiactoxicity, which method comprises administering to such a human in needthereof an amount of β-hydroxy-γ-trimethylaminobutyric acid, an isomerthereof, a pharmaceutically acceptable salt thereof, or apharmaceutically acceptable salt of an isomer thereof, sufficient toreduce cardiac toxicity.
 3. A method according to claim 2 wherein theβ-hydroxy-γ-trimethylaminobutyric acid is administered by ahydrochloride salt.